1. Field of the Invention
The present invention relates to a sensor utilizing attenuated total reflection (hereinafter referred to as ATR), such as a surface plasmon resonance sensor for quantitatively analyzing a substance in a sample by utilizing excitation of a surface plasmon, and more particularly to a sensor, utilizing ATR, of a type that detects a dark line occurring in a reflected light beam due to ATR by the use of photodetection means consisting of a plurality of light-receiving elements juxtaposed in a predetermined direction.
2. Description of the Related Art
In metals, if free electrons are caused to vibrate in a group, compression waves called plasma waves will be generated. The compression waves generated in a metal surface are quantized and called a surface plasmon.
A variety of surface plasmon resonance sensors have been proposed for quantitatively analyzing a substance in a sample by taking advantage of a phenomenon that a surface plasmon is exited by light waves. Among such sensors, one employing a system called xe2x80x9cKretschmann configurationxe2x80x9d is particularly well known (e.g., see Japanese Unexamined Patent Publication No. 6(1994)-167443).
The surface plasmon resonance sensor employing the xe2x80x9cKretschmann configurationxe2x80x9d is equipped mainly with a dielectric block formed, for example, into the shape of a prism; a metal film, formed on a surface of the dielectric block, for placing a sample thereon; a light source for emitting a light beam; an optical system for making the light beam enter the dielectric block at various angles of incidence so that the condition for total internal reflection is satisfied at the interface between the dielectric block and the metal film; and photodetection means for detecting the state of the surface plasmon resonance, that is, the state of ATR, by measuring the intensity of the light beam satisfying total internal reflection at the interface.
In order to obtain various angles of incidence, as described above, a relatively thin light beam may be caused to strike the above-mentioned interface at different angles of incidence, or relatively thick convergent or divergent rays may be caused to strike the interface so that they contain components incident at various angles. In the former, the light beam whose reflection angle varies with a change in the incidence angle of the incident light beam can be detected by a small photodetector that is moved in synchronization with the variation in the reflection angle, or by an area sensor extending in the direction in which the angle of reflection varies. In the latter, on the other hand, rays reflected at various angles can be detected by an area sensor extending in the direction in which all the reflected rays can be received.
In the surface plasmon resonance sensor mentioned above, if a light beam strikes the metal film at a specific incidence angle xcex8sp equal to or greater than a critical angle of incidence at which total internal reflection takes place, evanescent waves having electric field distribution are generated in the sample in contact with the metal film, whereby a surface plasmon is excited at the interface between the metal film and the sample. When the wave vector of the evanescent light is equal to the wave number of the surface plasmon and therefore the wave numbers between the two are matched, the evanescent waves and the surface plasmon resonate and light energy is transferred to the surface plasmon, whereby the intensity of light satisfying total internal reflection at the interface between the dielectric block and the metal film drops sharply. The sharp intensity drop is generally detected as a dark line by the above-mentioned photodetection means.
Note that the above-mentioned resonance occurs only when the incident light beam is a p-polarized light beam. Therefore, in order to make the resonance occur, it is necessary that a light beam be p-polarized before it strikes the interface.
If the wave number of the surface plasmon is found from an incidence angle xcex8sp at which ATR takes place, the dielectric constant of a sample can be obtained by the following Equation:
Ksp(xcfx89)=(xcfx89/c){∈m(xcfx89)∈s}1/2/{∈m(xcfx89)+∈s}1/2
where Ksp represents the wave number of the surface plasmon, xcfx89 represents the angular frequency of the surface plasmon, c represents the speed of light in vacuum, and ∈m and ∈s represent the dielectric constants of the metal and the sample, respectively.
If the dielectric constant ∈s of the sample is found, the density of a specific substance in the sample is found based on a predetermined calibration curve, etc. As a result, by finding the incidence angle xcex8sp at which the intensity of reflected light drops, the dielectric constant of the sample, that is, the properties of the sample related to the refractive index thereof can be specified.
In this kind of surface plasmon resonance sensor, photodetection means in the form of an array can be employed with the object of measuring the aforementioned incidence angle xcex8sp with a high degree of accuracy and in a large dynamic range, as disclosed in Japanese Unexamined Patent Publication No. 11(1999)-326194. The photodetection means is formed by a plurality of light-receiving elements juxtaposed in a predetermined direction. The light-receiving elements are disposed to respectively receive the components of a light beam satisfying total internal reflection at various angles of reflection at the aforementioned interface.
In that case, differentiation means is provided for differentiating the photodetection signals output by the light-receiving elements of the aforementioned photodetection means, in the direction where the light-receiving elements are juxtaposed. The properties of the sample related to the refractive index thereof are often analyzed based on differentiated values output by the differentiation means, particularly the differentiated value corresponding to a dark line that occurs in a reflected light beam.
In addition, a leaky mode sensor is known as a similar sensor making use of ATR, as disclosed, for instance, in xe2x80x9cSpectral Researches,xe2x80x9d Vol. 47, No. 1 (1998), pp. 21 to 23 and pp. 26 and 27. The leaky mode sensor is constructed mainly of a dielectric block in the form of a prism, for example; a cladding layer formed on a surface of the dielectric block; an optical waveguide layer, formed on the cladding layer, for placing a sample thereon; a light source for emitting a light beam; an optical system for making the light beam enter the dielectric block at various angles of incidence so that the condition for total internal reflection is satisfied at the interface between the dielectric block and the cladding layer; and photodetection means for detecting the excited state of the waveguide mode, that is, the state of ATR by measuring the intensity of the light beam satisfying total internal reflection at the interface between the dielectric block and the cladding layer.
In the leaky mode sensor with the construction mentioned above, if a light beam falls on the cladding layer through the dielectric block at angles of incidence equal to or greater than an angle of incidence at which total internal reflection takes place, the light beam is transmitted through the cladding layer and then only light with a specific wave number, incident at a specific angle, is propagated in the optical waveguide layer in a waveguide mode. If the waveguide mode is excited in this manner, the greater part of the incident light is confined within the optical waveguide layer, and consequently, ATR occurs in which the intensity of light satisfying total internal reflection at the above-mentioned interface drops sharply. Since the wave number of light propagating in the optical waveguide layer depends on the refractive index of the sample on the optical waveguide layer, the refractive index of the sample and/or the properties of the sample related to the refractive index thereof can be analyzed by finding the above-mentioned specific angle of incidence at which ATR takes place.
The leaky mode sensor also can employ the aforementioned photodetection means in the form of an array in order to detect the position of a dark line occurring in the reflected light by ATR. In addition, the aforementioned differentiation means is often employed along with the photodetection means.
In the field of pharmaceutical research, etc., the above-mentioned surface plasmon resonance sensor and leaky mode sensor are sometimes used in a random screening method of finding a specific substance that couples with a desired sensing medium. In this case, a sensing medium is placed on the aforementioned thin film layer (i.e., the metal film in the case of the surface plasmon resonance sensor, or the cladding layer and the optical waveguide layer in the case of the leaky mode sensor), and various solutions of substances (liquid sample) are added to the sensing medium, and each time a predetermined time elapses, the aforementioned differentiated value is measured. If the added substances are coupled with the sensing medium, the refractive index of the sensing medium varies with the lapse of time by the coupling. Therefore, by detecting the above-mentioned differentiated value every time a predetermined time elapses and then judging whether or not the differentiated value has been varied, it can be judged whether or not the added substances and the sensing medium have been coupled, that is, whether or not the added substances are specific substances that couple with the sensing medium. In this case, both the sensing medium and the liquid sample are samples to be analyzed. As such a combination of specific substances and a sensing medium, there is, for example, a combination of an antigen and an antibody.
A change in the differentiated value for each predetermined time, incidentally, is slight. Therefore, to measure a slight change in the differentiated value with a high degree of accuracy, it is desirable to amplify and detect the differentiated value with amplification means. However, the differentiated value corresponding to a dark line that occurs in a reflected light beam becomes a value corresponding to the positional relationship between the photodetection element and the dark line, and even if a differentiated value whose absolute value is smallest is selected, there is a fluctuation in the value. Because of this, in the case where the degree of amplification is great and the absolute value of the differentiated value is also great, there is a fear that subsequent electric circuits will be saturated. For this reason, sufficient amplification had not been able to be performed and measurements had not been able to be made with high sensitivity.
The present invention has been made in view of the circumstances mentioned above. Accordingly, it is the primary object of the present invention to provide a sensor, utilizing ATR, which is capable of accurately analyzing a sample by measuring a change with the lapse of time in a differentiated value with high sensitivity.
To achieve this end and in accordance with an important aspect of the present invention, there is provided a sensor utilizing attenuated total reflection, comprising:
a dielectric block;
a thin film layer, formed on a surface of the dielectric block, for placing a sample thereon;
a light source for emitting a light beam;
an optical system for making the light beam enter the dielectric block at various angles of incidence so that a condition for total internal reflection is satisfied at an interface between the dielectric block and the thin film layer;
photodetection means, comprising a plurality of light-receiving elements juxtaposed in a predetermined direction and disposed to respectively receive components of the light satisfying the total internal reflection condition at the interface, for detecting the attenuated total reflection;
differentiation means for differentiating a photodetection signal output from each of the light-receiving elements of the photodetection means, in the juxtaposed direction of the light-receiving elements, and then outputting a differentiated value; and
measurement means for subtracting an initial value from the differentiated value near a point where a change in the photodetection signal in the juxtaposed direction of the light-receiving elements makes a transition from decrease to increase, and then measuring a change with the lapse of time in the differentiated value from which the initial value has been subtracted.
In accordance with another important aspect of the present invention, there is provided a sensor utilizing attenuated total reflection, comprising:
a dielectric block;
a thin film layer, formed on a surface of the dielectric block, for placing a sample thereon;
a light source for emitting a light beam;
an optical system for making the light beam enter the dielectric block at various angles of incidence so that a condition for total internal reflection is satisfied at an interface between the dielectric block and the thin film layer;
photodetection means, comprising a plurality of light-receiving elements juxtaposed in a predetermined direction and disposed to respectively receive components of the light satisfying the total internal reflection condition at the interface, for detecting the attenuated total reflection caused by surface plasmon resonance;
differentiation means for differentiating a photodetection signal output from each of the light-receiving elements of the photodetection means, in the juxtaposed direction of the light-receiving elements, and then outputting a differentiated value; and
measurement means for subtracting an initial value from the differentiated value near a point where a change in the photodetection signal in the juxtaposed direction of the light-receiving elements makes a transition from decrease to increase, and then measuring a change with the lapse of time in the differentiated value from which the initial value has been subtracted.
In accordance with still another important aspect of the present invention, there is provided a sensor utilizing attenuated total reflection, comprising:
a dielectric block;
a cladding layer formed on a surface of the dielectric block;
an optical waveguide layer, formed on a surface of the cladding layer, for placing a sample thereon;
a light source for emitting a light beam;
an optical system for making the light beam enter the dielectric block at various angles of incidence so that a condition for total internal reflection is satisfied at an interface between the dielectric block and the thin film layer;
photodetection means, comprising a plurality of light-receiving elements juxtaposed in a predetermined direction and disposed to respectively receive components of the light satisfying the total internal reflection condition at the interface, for detecting the attenuated total reflection caused by excitation of a waveguide mode in the optical waveguide layer;
differentiation means for differentiating a photodetection signal output from each of the light-receiving elements of the photodetection means, in the juxtaposed direction of the light-receiving elements, and then outputting a differentiated value; and
measurement means for subtracting an initial value from the differentiated value near a point where a change in the photodetection signal in the juxtaposed direction of the light-receiving elements makes a transition from decrease to increase, and then measuring a change with the lapse of time in the differentiated value from which the initial value has been subtracted.
In the aforementioned sensors utilizing attenuated total reflection (ATR), it is preferable that the xe2x80x9cdifferentiated value near a point where a change in the photodetection signal makes a transition from decrease to increasexe2x80x9d be a differentiated value closest to a point where a change in the photodetection signal makes a transition from decrease to increase. However, the present invention is not limited to the differentiated value closest to the point. The differentiated value may be a differentiated value near a point where a change in the photodetection signal makes a transition from decrease to increase. The aforementioned xe2x80x9cinitial valuexe2x80x9d may employ the value of a differentiated value near a point where a change in the photodetection signal makes a transition from decrease to increase. Alternatively, by performing an arithmetic process such as feedback at the time of the start of measurements and then finding a value which shifts a measured value at the time of the start of measurements to the vicinity of 0, it may be set as an initial value. In doing subtraction, an initial value is subtracted from the differentiated value near a point where a change in the photodetection signal makes a transition from decrease to increase, by the use of a subtracter, etc. Furthermore, subtraction may be done by finding a difference between the two with a differential circuit, etc.
In the sensor of the present invention, it is preferable that the differentiation means output a difference between the optical signals output from two adjacent light-receiving elements of the photodetection means. It is also preferable that the photodetection means be a photodiode array.
In the sensor of the present invention, the aforementioned differentiation means can employ an analog circuit. In addition, the aforementioned measurement means may perform the subtraction by an analog circuit, and after A/D conversion is performed on the differentiated value from which the initial value has been subtracted, the measurement is made by a digital circuit. The aforementioned measurement means may further comprise amplification means comprising an analog circuit for amplifying the differentiated value from which the initial value has been subtracted.
In addition, The sensor of the present invention utilizing ATR may further comprise filtration means for performing a filtration process on the differentiated value.
The expression xe2x80x9cperforming a filtration process on the differentiated value,xe2x80x9d in addition to performing the filtration process on the differentiated value itself, includes performing the filtration process on the photodetection signals that are used for calculating the differentiated value, performing the filtration process on the differentiated value from which an initial value has been subtracted, and so on.
In the sensor of the present invention, the filtration means can employ, for example, a low-pass filter that allows a signal of frequency 100 Hz or less to pass through it. The low-pass filter may be a filter that allows a signal of frequency 10 Hz or less to pass through it.
According to the sensor of the present invention utilizing ATR, the photodetection means in the form of an array, consisting of a plurality of light-receiving elements juxtaposed, is employed and the photodetection signals output from the light-receiving elements are differentiated in the juxtaposed direction of the light-receiving elements by differentiation means. A change with the lapse of time in the differentiated value is measured to analyze the properties of a sample. Because of this, the initial value of a differentiated value is first measured. Every time a differentiated value is measured, the initial value is subtracted from the differentiated value output from the differentiation means. As a result, the differentiated value from which the initial value has been subtracted contains no fluctuation in an absolute value corresponding to the positional relationship between the photodetector element and a dark line, and becomes a value reflecting only a change with the lapse of time in the differentiated value from the time of the start of measurements. Therefore, the value after the subtraction is small in absolute value, compared with a differentiated value on which such subtraction is not done, and can be amplified by a sufficiently high amplification factor. Thus, a change with the lapse of time in a differentiated value can be measured with high sensitivity and an accurate analysis of a sample can be made.